In Vitro Microtubule Dynamics Assays Using Dark-Field Microscopy.
In Vitro Microtubule Dynamics Assays Using Dark-Field Microscopy.
复制标题
使用暗场显微镜进行体外微管动力学测定。
DOI：
10.1007/978-1-0716-0219-5_4
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Roll-Mecak,Antonina
中科院分区：
文献类型：
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作者：
Spector,JeffreyO;Vemu,Annapurna;Roll-Mecak,Antonina
微管是动态的非共价介观聚合物。它们的动态行为对细胞生物学过程至关重要，从细胞内运输到细胞分裂和神经发生。在过去的二十年中，荧光显微镜一直是监测微管动力学的首选方法。然而，荧光微管容易受到光损伤，从而改变其动力学，荧光标签本身也会影响微管的性质。暗场成像是一种无标记技术，可以产生高信噪比、低背景的微管图像，以高采集率，没有荧光显微镜固有的光漂白。在这里，我们描述了如何成像体外微管动力学使用暗场显微镜。无标记微管成像的能力允许对荧光标记不可行的非丰富微管物种的动态特性进行调查，避免了添加荧光标记引起的混淆效应。
Microtubules are dynamic non-covalent mesoscopic polymers. Their dynamic behavior is essential for cell biological processes ranging from intracellular transport to cell division and neurogenesis. Fluorescence microscopy has been the method of choice for monitoring microtubule dynamics in the last two decades. However, fluorescent microtubules are prone to photodamage that alters their dynamics, and the fluorescent label itself can affect microtubule properties. Dark-field imaging is a label-free technique that can generate high signal-to-noise, low-background images of microtubules at high acquisition rates without the photobleaching inherent to fluorescence microscopy. Here, we describe how to image in vitro microtubule dynamics using dark-field microscopy. The ability to image microtubules label-free allows the investigation of the dynamic properties of non-abundant tubulin species where fluorescent labeling is not feasible, free from the confounding effects arising from the addition of fluorescent labels.